Calcium Ionophore, Calcimycin, Kills Promastigotes by Activating Parasite Nitric Oxide Synthase.

Leishmaniasis is an infectious disease caused by protozoan parasites of the genus . There is no vaccine against human leishmaniasis and the treatment of the disease would benefit from a broader spectrum and a higher efficacy of leishmanicidal compounds. We analyzed the leishmanicidal activity and the mechanism of action of the calcium ionophore, calcimycin. promastigotes were coincubated with calcimycin and the viability of the cells was assessed using resazurin assay. Calcimycin displayed dose-dependent effect with IC = 0.16 M. Analysis of propidium iodide/LDS-751 stained promastigotes revealed that lower concentrations of calcimycin had cytostatic effect and higher concentrations had cytotoxic effect. To establish the mechanism of action of calcimycin, which is known to stimulate activity of mammalian constitutive nitric oxide synthase (NOS), we coincubated promastigotes with calcimycin and selective NOS inhibitors ARL-17477 or L-NNA. Addition of these inhibitors substantially decreased the toxicity of calcimycin to promastigotes. In doing so, we demonstrated for the first time that calcimycin has a direct leishmanicidal effect on promastigotes. Also, we showed that constitutive Ca/calmodulin-dependent nitric oxide synthase is involved in the parasite cell death. These data suggest activation of nitric oxide synthase as a new therapeutic approach.